Selectivity and engineering of the sialoglycan-binding spectrum in Siglec-like adhesins

Abstract

The Siglec-like Serine-Rich Repeat (SRR) adhesins mediate bacterial attachment to mammalian hosts via sialoglycan receptors. Here, we combine structural, computational, biochemical, and phylogenetic approaches to elucidate the determinants of the sialoglycan-binding spectrum across the family of Siglec-like SRR adhesins. We further identified mutable positions that disproportionately affect sialoglycan selectivity, as demonstrated by increases in binding to alternative ligands of 2-to 3-orders of magnitude. Biologically, these studies highlight how bacteria nimbly modulate the receptor interaction during coevolution of host and pathogen. These studies additionally created binding proteins specific for sialyl-T antigen or 6S-sialyl Lewis X that can recognize glycosylation of human plasma proteins. The engineered binding proteins can facilitate the characterization of normal cellular glycan modifications or may be used as diagnostic tools in disease states with altered glycosylation.Significance: The ability of bacteria to bind selectively to host receptors underlies both commensalism and pathogenesis. Here, we identify the molecular basis for receptor selectivity in streptococci that bind to sialoglycan receptors. This revealed how to convert these adhesins into selective probes that measure tri-and tetrasacharides within the context of larger glycosylations. These probes that can be used in a laboratory with no specialized equipment and can be used to address biological questions relating to sialoglycan-dependent signaling and adhesion.

Tandem sialoglycan-binding modules in a Streptococcus sanguinis serine-rich repeat adhesin create target dependent avidity effects

“… SK1 interactions with ligands. The structures of the Siglec domains of A–B , SK1 Siglec1+Unique1 , C–D , SK1 Siglec2+Unique2 , and E , Hsa (PDB entry 6EFD ) ( 18 ) are shown in cartoon in teal , lavender , and orange , respectively. The adhesin residues that hydrogen bond with the ligands are shown as sticks .…”

“…As compared with adhesins with canonical YTRY motifs ( 16 , 18 , 19 ), the sialic acid of each trisaccharide interacts with the two noncanonical motifs of SK1 via fewer hydrogen-bonding interactions ( Fig. 6 , A–E ).…”

“…Interaction of the S. gordonii and S. sanguinis SRR adhesins with host sialoglycan structures relies upon a domain within the binding region related in fold to mammalian s ialic acid–binding i mmuno g lobulin-like lec tins (Siglecs); indeed both are organized around a V-set Ig fold ( 6 , 16 – 18 ). These “Siglec-like” SRR adhesins always contain a second domain immediately following the Siglec domain ( 16 ).…”

“…Recent structural and engineering studies revealed that sialoglycan binding and selectivity are also affected by three adjacent loops of high sequence diversity ( 18 ). Using nomenclature from the V-set Ig fold identified them as the CD loop, the EF loop, and the FG loop ( 16 ).…”

“…Amino acid side chains in these loops directly hydrogen-bond with sialoglycan ligands ( 16 , 17 , 19 ), and the sequence diversity of these loops is proposed as a major determinant of sialoglycan selectivity in the Siglec-like adhesins. As a result, they have been termed “selectivity loops” ( 18 ).…”